Optimization of biohydrogen production from sweet sorghum syrup using statistical methods

This study employed statistically based experimental designs to optimize fermentation conditions for hydrogen production from sweet sorghum syrup by anaerobic mixed cultures. Initial screening of important factors influencing hydrogen production, i.e., total sugar, initial pH, nutrient solution, iron (II) sulphate (FeSO₄), peptone and sodium bicarbonate was conducted by the Plackett–Burman method. Results indicated that only FeSO₄ had statistically significant (P ≤ 0.005) influences on specific hydrogen production (P_s) while total sugar and initial pH had an interdependent effect on P_s. Optimal conditions for the maximal P_s were 25 g/L total sugar, 4.75 initial pH and 1.45 g/L FeSO₄ in which P_s of 6897 mL H₂/L was estimated. Estimated optimum conditions revealed only 0.04% difference from the actual P_s of 6864 mL H₂/L which suggested that the optimal conditions obtained can be practically applied to produce hydrogen from sweet sorghum syrup with the least error.     

Optimization of biohydrogen production by the novel psychrophilic strain N92 collected from the Antarctica

In this study, the response surface methodology (RSM) with central composite design (CCD) was employed to improve the hydrogen production by the psychrophilic N92 strain (EU636058) isolated from Antarctica, which is closely related to Pseudorhodobacter sp. (KT163920). The influence of operational conditions such as temperature (4.7–55.2 °C), initial pH (3.44–10.16), and initial glucose concentration (4.7–55.23 g/dm³), as well as the initial concentrations of (NH₄)₂SO₄ (0.05–3.98 g/dm³), FeSO₄ (0.02–1.33 g/dm³) and NaHCO₃ (0.02–3.95 g/dm³) was evaluated. The linear effect of glucose concentration, along with the quadratic effect of all the six factors were the most significant terms affecting the biohydrogen yield by N92 strain. The optimum conditions for the maximum hydrogen yield of 1.7 mol H₂/mol glucose were initial pH of 6.86, glucose concentration of 28.4 g/dm³, temperature 29 °C and initial concentration of (NH₄)₂SO₄, FeSO₄ and NaHCO₃ of 0.53, 1.55 and 1.64 g/dm³ respectively. Analysis of the metabolites produced under the optimum conditions showed that the most abundant were acetic acid (0.8 g/dm³), butyric acid (0.7 g/dm³) and ethanol (2.1 g/dm³). We suggest that the bioprocess established in this study using the strain N92 could be an alternative for hydrogen production with the advantages of constituting low energy costs in fermentation.     

Optimization of culture conditions for biohydrogen production from sago wastewater by Enterobacter aerogenes using Response Surface Methodology

Sago wastewater (SWW) causes pollution to the environment due to its high organic content. Annually, about 2.5 million tons of SWW is produced in Malaysia. In this study, the potential of SWW as a substrate for biohydrogen production by Enterobacter aerogenes (E. aerogenes) was evaluated. Response Surface Methodology (RSM) was employed to find the optimum conditions. From preliminary optimization, it was found that the most significant factors were yeast extract, temperature, and inoculum size. According to Face Centered Central Composite Design (FCCCD), the maximum hydrogen concentration and yield were 630.67 μmol/L and 7.42 mmol H₂/mol glucose, respectively, which is obtained from the sample supplemented with 4.8 g/L yeast extract concentration, 5% inoculum, and incubated at the temperature of 31 °C. Cumulative hydrogen production curve fitted by the modified Gompertz equation suggested that H_max, R_max, and λ from this study were 15.10 mL, 2.18 mL/h, and 9.84 h, respectively.     

Optimization of biohydrogen inoculum development via a hybrid pH and microwave treatment technique – Semi pilot scale production assessment

The interactive effect of a hybrid pH and microwave pre-treatment on a mixed inoculum for biohydrogen production was investigated. Response surface methodology (RSM) was employed to obtain the optimum pre-treatment conditions of pH, microwave duration and microwave intensity for maximum hydrogen yield. The obtained model had a coefficient of correlation (R²) of 0.87. The optimum inoculum pre-treatment conditions predicted were pH 11 and 2 min microwave treatment at 860 W and the validation experiments demonstrated a 32.41% increase on hydrogen yield.     

Cladophora sp. as a sustainable feedstock for dark fermentative biohydrogen production

Macroalgae are rich in carbohydrates which can be used as a promising substrate for fermentative biohydrogen production. In this study, Cladophora sp. biomass was fermented for biohydrogen production at various inoculum/substrate (I/S) ratios against a control of inoculum without substrate in laboratory-scale batch reactors. The biohydrogen production yield ranged from 40.8 to 54.7 ml H₂/g-VS, with the I/S ratio ranging from 0.0625 to 4. The results indicated that low I/S ratios caused the overloaded accumulation of metabolic products and a significant pH decrease, which negatively affected hydrogen production bacteria's metabolic activity, thus leading to the decrease of hydrogen fermentation efficiency. The overall results demonstrated that Cladophora sp. biomass is an efficient fermentation feedstock for biohydrogen production.     

Modelling and optimization of xylose and glucose production from napier grass using hybrid pre-treatment techniques

This work models and optimizes four hybrid techniques of Napier grass pre-treatment for xylose and glucose production, namely HCl and moist heat (HH), HCl and microwave (HM), NaOH and moist heat (NH) and NaOH and microwave (NM) using the Response Surface Methodology (RSM). The coefficients of determination (R²) of 0.83 and 0.97 were obtained for xylose and glucose production respectively using HH hybrid pre-treatment, and 0.90 and 0.80 were obtained for xylose and glucose respectively using HM hybrid pre-treatment. The optimized pre-treatment conditions of HH gave 12.83 gL⁻¹ xylose and 2.28 gL⁻¹ glucose, and optimized HM pre-treatment gave 15.06 g L⁻¹ and 2.44 gL⁻¹ xylose and glucose respectively. A xylose to glucose concentration ratio of 5.6:1 was obtained for the optimized HH pre-treatment compared to 6.1:1 for the optimized HM pre-treatment. For NH and NM hybrid pre-treatments, low concentrations of fermentable sugars were observed (<0.5 gL⁻¹). The findings indicate that xylose and glucose production from Napier grass can be enhanced by an optimal combination of pre-treatments of HCl and moist heat at a volume fraction of 4.39% HCl, 93.07 °C for 180 min, or using a combination of microwave and HCl at a volume fraction of 5% HCl, 500 W for 30 min.     

Optimization for simultaneous enhancement of biobutanol and biohydrogen production

Acetone-butanol-ethanol (ABE) fermentation guarantees a sustainable route for biohydrogen and biobutanol production. This research work is committed towards the enhancement of biohydrogen and biobutanol production by single and multi-parameter optimization for the improvement of substrate energy recovery using C. saccharoperbutylacetonicum. Single parameters optimization (SPO) manifested that headspace of 60% (v/v) and butyric acid supplementation of 9 g/L and temperatures of 30 °C and 37 °C were suitable for obtaining maximum biohydrogen and biobutanol production, respectively. The interaction between these parameters was further evaluated by implementing a 5-level 3-factor Central Composite Design (CCD). In the present study, a central composite design was employed to enhance the biohydrogen and biobutanol production. In addition, the experimental results were analyzed by response surface methodology (RSM) and artificial intelligence (AI) techniques such as artificial neural network (ANN). The prediction capability of RSM was further compared with ANN for predicting the optimum parameters that would lead to maximum biohydrogen and biobutanol production. ANN yielded higher values of biohydrogen and biobutanol. ANN was found to be superior as compared to RSM in terms of prediction accuracy for both biohydrogen and biobutanol because of its higher coefficient of determination (R²) and lower root mean square error (RMSE) value. Process temperature (32.65 °C), headspace (58.21% (v/v)) and butyric acid supplementation (9.16 g/L) led to maximum substrate energy recovery of 78% with biohydrogen and biobutanol production of 5.9 L/L and 16.75 g/L, respectively. Process parameter optimization led to a significant increase in substrate energy recovery from Biphasic fermentation.     

Optimization of conditions for hydrogen production from complex dairy wastewater by anaerobic sludge using desirability function approach

Present study deals with the multiple-response optimization for biohydrogen production using anaerobic sludge and outstanding approach to overcome the drawbacks of conventional response surface methodology (RSM). Dairy wastewater was used as source in batch fermentation was followed for this study. Response surface methodology (RSM), based on a three level, four variable Box–Behnken design, was employed to obtain the best possible combination of substrate concentration, pH, COD/N ratio and COD/P ratio for maximum H₂ yield (HY) and specific hydrogen production rate (SHPR). Experimental data were evaluated by applying RSM integrating a desirability function approach. The optimum H₂ yield and SHPR conditions were: substrate concentration 15.3 g COD/L, pH 5.5, COD/N ratio 100.5 and COD/P ratio 120 with maximum overall desirability D of 0.94. The confirmation experiment under these optimal condition showed a HY and SHPR of 13.54 mmol H₂/g COD and 29.91 mmol H₂/g-VSS.d, respectively. This was only 0.22% and 0.20%, respectively, different from the predicted values, suggesting that the desirability function approach with RSM was a useful technique to get the maximum H₂ yield and SHPR simultaneously.     

Impact of pretreatment on food waste for biohydrogen production: A review

Food waste (FW) can be utilized as a raw material to produce energy such as hydrogen via fermentation, which is a more attractive and environmentally friendly approach compared to incineration and land-filling. Food waste must be pretreated before being used in various biological processes. The choice of the pretreatment method usually depends on the composition of the food waste. Therefore, various pretreatment methods generally employed to treat FW, including physical, physiochemical, chemical and biological pretreatments, are summarized in this review. The different pretreatment methods are compared in terms of their efficiency and biohydrogen yield. Additionally, the energy efficiencies of the various pretreatment methods are compared, thereby leading to the selection of the most efficient pretreatment method.     

Production and optimization of high grade cellulase from waste date seeds by Cellulomonas uda NCIM 2353 for biohydrogen production

Production of high grade cellulolytic enzymes from waste agricultural biomass would valorise these wastes to valuable products as well as avoid the pollution problems associated with landfilling of the biomass. In the present study, waste date palm (Phoenix dactylifera) seeds were valorised for cellulase production from Cellulomonas uda NCIM 2353 and for its subsequent usage in biohydrogen production. Optimization of key operational parameters such as date seed concentration, xylose, casein and initial media pH were performed using central composite design to obtain the maximum enzyme yield. The optimum values obtained were (g/L): date seed concentration 30.65, xylose concentration 0.55, casein 7.00 and pH 7.40 for a determination coefficient of 0.999. The results demonstrated a higher prediction accuracy of response surface methodology as the cellulase activity increased six fold (175.96 IU/mL) after optimization. The optimum pH and temperature of purified cellulase was 7 and 50 °C respectively where the enzyme retained nearly 80% of activity upto 180 min. Enzymatic hydrolysis studies showed that a high saccharification efficiency of 60.5% was obtained for acid pretreated sugarcane bagasse by the indigenous cellulase, equivalent to the performance of commercial cellulase. Further, the as-obtained reducing sugars were decomposed by Clostridium thermocellum to produce biohydrogen of maximum concentration 187.44 mmol/L at end of 24 h of fermentation. Results show that date seed substrate based cellulase protein can be employed for industrial processes of biohydrogen production.     

Optimization of particle number,substrate concentration and temperature of batch immobilized reactor system for biohydrogen production by dark fermentation

Immobilized cell bioreactor was operated in batch mode for biohydrogen generation by dark fermentation from acid hydrolyzed waste wheat powder. It was aimed to optimize the fermentation conditions with the purpose of obtaining the highest hydrogen yield (Y_H2) and production rate (HPR) by applying Box–Wilson statistical experimental design method. Particle number (PN = 120–240; X₁), initial total sugar concentration (TS₀ = 10–30 g/l; X₂) and fermentation temperature (T = 35–55 °C; X₃) were selected as independent variables. Polyester fibers with particle diameter “Dp” = 0.5 cm were used as support material to immobilize microorganisms with heat-pretreated sludge. Quadratic equations for production yield and rate were developed by using experimental results. The maximum Y_H2 (3.21 mol H₂/mol glucose) and HPR (73.3 ml H₂/h) were predicted at the optimum conditions of PN = 240, TS₀ = 10 g/l and T = 44.9 °C. Also, analysis of variance, as well as sum of ranking difference test results demonstrated that fitting models were statistically significant.     

A review on process modeling and design of biohydrogen

The current energy supply depends on fossil fuels which have increased carbon dioxide emissions leading to global warming and depleted non-renewable fossil fuels resources. Hydrogen (H₂) fuel could be an eco-friendly alternative since H₂ consumption only produces water. However, the overall impacts of the H₂ economy depend on feedstock types, production technologies, and process routes. The existing process technologies for H₂ production used fossil fuels encounter the escalation of fossil fuel prices and long-term sustainability challenges. Therefore, biohydrogen production from renewable resources like biomass wastes and wastewaters has become the focal development of a sustainable global energy supply. Different from other biohydrogen production studies, this paper emphasizes biohydrogen fermentation processes using different renewable sources and microorganisms. Moreover, it gives an overview of the latest advancing research in different biohydrogen process designs, modeling, and optimization. It also presents the biohydrogen production routes and kinetic modeling for biohydrogenation.     

Optimization of fermentative hydrogen production by Enterococcus faecium INET2 using response surface methodology

A newly isolated strain Enterococcus faecium INET2 was used as inoculum for biohydrogen production through dark fermentation. The individual and interactive effect of initial pH, operation temperature, glucose concentration and inoculation amount on the accumulation of hydrogen during fermentation was examined by a Box–Behnken Design (BBD), and hydrogen production process was analyzed at the optimal condition. A significant interactive effect between glucose concentration and pH was observed, the optimal condition was initial pH 7.1, operation temperature 34.8 °C, glucose concentration 11.3 g/L and inoculation amount 10.4%. Hydrogen yield, maximum hydrogen production rate and hydrogen production potential were determined to be 1.29 mol H₂/mol glucose, 86.7 L H₂/L/h and 1.35 L H₂/L. Metabolites analysis showed that E. faecium INET2 followed the pyruvate: formate lyase (Pfl) pathway in first 16 h, followed by the acetate-type fermentation and then shifted to butyrate-type fermentation. Maximum hydrogen production rate was accompanied with a quick formation of acetic acid.     

Continuous biohydrogen production by a degenerated strain of Clostridium acetobutylicum ATCC 824

Degenerated strains of Clostridium acetobutylicum lack the ability to produce solvents and to sporulate, allowing the continuous production of hydrogen and organic acids. A degenerated strain of Clostridium acetobutylicum was obtained through successive batch cultures. Its kinetic characterization showed a similar specific growth rate than the wild type (0.25 h^?1), a higher butyric acid production of 6.8 g·L^?1 and no solvents production. A steady state was reached in a continuous culture at a dilution rate of 0.1 h^?1, with a constant hydrogen production of 507 mL·h^?1, corresponding to a volumetric rate of 6.10 L·L^?1 d^?1, and a yield of 2.39 mol of H₂ per mole of glucose which represents 60% of the theoretical maximum yield. These results suggest that the degeneration is an interesting alternative for hydrogen production with this strain, obtaining a high hydrogen production in a continuous culture with cells in a permanent acidogenic state.     

Multiscale mixing analysis and modeling of biohydrogen production by dark fermentation

Hydrogen production by dark fermentation (DF) from wastewater, food waste, and agro-industrial waste combines the advantages to be renewable, sustainable and environmentally friendly. But this attractive process involves a three-phase gas-liquid-solid system highly sensitive to mixing conditions. However, mixing is usually disregarded in the conventional strategies for enhancing biohydrogen productivity, even though H₂ production can be doubled, e.g. versus of reactor design (0.6–1.5 mol H₂/mol hexose). The objective of this review paper is, therefore, to highlight the key effects of mixing on biohydrogen production among the abiotic parameters of DF. First, the pros and cons of the different modes of mixing in anaerobic digesters are described. Then, the influence of mixing on DF is discussed using recent data from the literature and theoretical analysis, focusing on the multiphase and multiscale aspects of DF. The methods and tools available to quantify experimentally the role of mixing both at the local and global scales are summarized. The 0-D to 3-D strategies able to implement mixing in fermentation modeling and scale-up procedures are examined. Finally, the perspectives in terms of process intensification and scale-up tools using mixing optimization are discussed with the issues that are still to be solved.     

Optimization of key factors affecting hydrogen production from food waste by anaerobic mixed cultures

Key factors (inoculums concentration, substrate concentration and citrate buffer concentration) affecting hydrogen yield (HY) and specific hydrogen production rate (SHPR) from food waste in batch fermentation by anaerobic mixed cultures were optimized using Response Surface Methodology with Central Composite Design. The experiments were conducted in 120 ml serum bottles with a working volume of 70 mL. Under the optimal condition of 2.30 g-VSS/L of inoculums concentration, 2.54 g-VS/L of substrate concentration, and 0.11 M of citrate buffer concentration, the predicted maximum HY and SHPR of 104.79 mL H₂/g-VS_added and 16.90 mL H₂/g-VSS.h, respectively, were obtained. Concentrations of inoculums, substrate and citrate buffer all had an individual effect on HY and SHPR (P < 0.05). The substrate concentration and citrate buffer concentration had the greatest interactive effect on SHPR (P = 0.0075) while their effects on HY (P = 0.0131) were profound. These results were reproduced in confirmation experiments under optimal conditions and generated an HY of 104.58 mL H₂/g-VS_added and an SHPR of 16.86 mL H₂/g-VSS.h. This was only 0.20% and 0.24%, respectively, different from the predicted values. Microbial community analysis by PCR-DGGE indicated that Clostridium was the pre-dominant hydrogen producer at the optimum and worst conditions. The presence of Lactobacillus sp. and Enterococcus sp. might be responsible for the low HY and SHPR at the worst condition.     

Modeling of biohydrogen production using generalized multi-scale kinetic model: Impacts of fermentation conditions

This paper presents a new multi-scale kinetic model built upon the multi-stage growth Hypothesis for predicting biohydrogen production. The proposed model represents the significant factors affecting biohydrogen production using a sum of first-order kinetic terms with varying dynamics from slow to fast one. The current work investigates 52 case studies of biohydrogen production that show the double first-order kinetic model provides the best modeling fitness (R² > 0.99). This result suggests two prevalent pathways or microbial groups with distinct dynamics (i.e., fast and slow modes) in biohydrogen production. An increase in temperature (30 °C–43 °C) or substrate concentration (10 g/L to 40 g/L) and the use of simple substrates or mixed cultures can increase the fast-mode dominance up to 100% contribution. Model analysis suggests that the fast mode corresponds to the butyrate production pathway, the growth-associated hydrogen-producing activity, the easily-biodegradable substrates, or the quick hydrogen-producing groups.     

A comprehensive review on biohydrogen production pilot scale reactor technologies: Sustainable development and future prospects

The current study focuses on a comprehensive review of the pilot scale production of biohydrogen and various factors affecting the design experiments. Biohydrogen is a clean energy carrier that can be used as a potential alternative to fossil fuels. Biohydrogen as a fuel has several advantageous attributes, including; carbon-neutral or carbon-zero nature, easy renewability, eco-efficient productivity (via biological routes), eco-friendly conversion, and the highest energy content among all existing fuels. Pilot-scale production of biohydrogen is limited because it requires a better understanding of the possible interactions involved in the process. In this review, biohydrogen production on various types of reactors such as stirred tank reactors, packed bed reactors, fluidized bed reactors, trickling filter reactors, etc., have been discussed. However, biohydrogen production has been mostly studied on small scale, the most challenging issue concerning large-scale production of biohydrogen is its relatively high cost over fuels from fossil owing to high feedstock and manufacturing costs. Therefore, cost-effective and eco-friendly biohydrogen production technologies should be necessarily developed and continuously improved to make this biofuel more competitive over its counterpart. In comparison with fossil fuels, biohydrogen has a high energy yield and is highly renewable. It can fulfill the future demand as a transport fuel.     

Production of biohydrogen from hydrolyzed bagasse with thermally preheated sludge

Production of biohydrogen from dark fermentation is an interesting alternative to producing renewable fuels because of its low cost and various usable substrates. Cellulosic content in plentiful bagasse residue is an economically feasible feedstock for biohydrogen production. A statistical experimental design was applied to identify the optimal condition for biohydrogen production from enzymatically hydrolyzed bagasse with 60-min preheated seed sludge. The bagasse substrate was first heated at 100 °C for 2 h and was then hydrolyzed with cellulase. Culture of the pretreated bagasse at 55 °C provided a higher H₂ production performance than that obtained from cultures at 45 °C, 65 °C, 35 °C and 25 °C. On the other hand, the culture at pH 5 resulted in higher H₂ production than the cultures at pH 6, pH 4 and pH 7. The optimal culture condition for the hydrogen production rate was around 56.5 °C and pH 5.2, which was identified using response surface methodology. Moreover, the pretreatment of bagasse under alkaline conditions gave a thirteen-fold increase in H₂ production yield when compared with that from preheatment under neutral condition.     

Recent insights into consolidated bioprocessing for lignocellulosic biohydrogen production

Biohydrogen production via dark fermentation using fermentable sugars from biomass materials is a sustainable way of procuring biohydrogen. Lignocellulosic biomass is a potential renewable feedstock for dark fermentation, but its use is challenged by the recalcitrant nature and generation of certain fermentation inhibitors resulting in compromised fermentation performance. Consolidated bioprocessing (CBP), the successful integration of hydrolysis and fermentation of lignocellulosic biomass to desirable products, has received tremendous research attentions in recent years to boost renewable fuel production in an economically feasible way. A microbial strain capable of both biomass hydrolysis and hydrogen fermentation is critical for successful CBP-based hydrogen fermentation. This review provides comprehensive information on dark fermentation for hydrogen production using lignocellulosic biomass as a potential feedstock with a CBP approach. Consolidated bioprocessing of lignocellulosic biomass for biohydrogen production via native and recombinant microbial strains is discussed in detail. Potential bottlenecks in the above mentioned processes are critically analyzed and future research perspectives are presented.